Studies in this project have shown that mouse oocytes actively regulate gene expression in specific populations of granulosa cells, and orchestrate the overall rate and organization of follicular development. The development of both oocytes and companion follicular cells appears to be governed by an oocyte- granulosa cell regulatory loop. The objectives of this renewal proposal are to (Part A) define roles of the oocyte in establishing patterns of gene expression in granulosa cells and (Part B) identify and functionally characterize key oocyte components of the oocyte-granulosa cell regulatory loop. Part A. The transition from preantral to antral follicle is a key stage of follicular development when the oocyte undergoes critical developmental transitions: oocytes become competent to resume meiosis and begin to acquire competence to undergo fertilization and embryogenesis. In addition, granulosa cells become divided into two groups during this transition: granulosa cells surrounding the oocyte become a morphologically and functionally distinct population called cumulus cells from those lining the follicle wall (mural granulosa cells). In Specific Aim 1, marker genes uniquely expressed by cumulus cells of preovulatory follicles will be identified. Then, the hypothesis that expression of these marker genes is regulated by oocytes will be tested. Part B. A yeast- based signal sequence trap (SST) was used to identify secreted and transmembrane proteins produced by mouse oocytes that could function in the oocyte-granulosa cell regulatory loop. The SST identified both known and novel proteins. Specific Aim 2 tests the hypothesis that one of the known proteins, CRUMBS, is involved in the organization of the granulosa cell epithelium, particularly during the preantral to antral follicle transition. This hypothesis will be tested by characterizing the follicular phenotype of an induced null mutation produced by targeted disruption of the Crbl gene. Specific Aim 3 tests the hypothesis that novel proteins identified by the SST strategy and expressed only by oocytes are key regulators of follicular development. The expression of genes encoding these proteins will be determined throughout follicular development and a functional analysis will be conducted by producing knockout models with null mutations. These experiments will identify key components of the regulatory loop and determine how they function to coordinate and drive the development of the oocyte-granulosa cell complex. Defects in the loop result in infertility, and the proteins participating in it might be targets for fertility control.